Fusion gene of non-essential glycoprotein gG (gX) and lacZ was constructed and pseudorabies virus (PrV) mutants were produced. Characteristics and usefulness of this mutant virus as a transneuronal tracer was investigated by injecting it into masseter or stomach of SD rats. Injection of PrV solution with titer of 1.0*10^7p.f.u./ml was most suitable for the tracing study. Injection of higher titer of viral solution killed the animals within 2 days, indicating the retained virulence of this mutant virus. Although viral infection occurred in motor trigeminal n.neurous in all cases of masseter injection, labeled neurous in the mesencephalic trigeminal n.were very few. Second-or third-order neurons were identified in the locus coeruleus (LC), raphe magnus (RM), medullary reticular formation (IRt), Edinger-Westphal nucleus (EW), tuberal magnocellular n.(TM), lateral hypothalamic n.(LH) and paraventricular hypothalamic n.(PVH). No anterograde tracing was detected in the CNS.Injection of this virus into stomach labeled neurons in the dorsal motor nucleus of N.X (DMN), n.tractus solitarius, n. ambiguus, A5 and PVH.These findings indicate that this virus preferentially infecct somatic and visceral motoneurons and is retrogradely tranported in second-order or third-order neurons in the CNS.